PLATING TECHNIQUES

When performing Antimicrobial Textile Testing, different plating techniques are used with the purpose of isolating and/or enumerating bacteria. The two main techniques used will be explained.

STREAK PLATE

In the streak plate method, the bacterial solution is spread over the surface of an agar plate in separated points so that, after incubation, single bacterial colonies can be isolated. During Antimicrobial Textile Testing this is the preferred method to use when preparing the stock culture plate.

The spread of the bacterial solution on the agar´s surface can be done following different patterns. This is one of the available procedures to performing streak plate:

INSTRUCTIONS

  1. Immerse the cotton swab into the bacterial solution
  2. Gently roll the cotton swab horizontally onto 1 / 3 of the agar´s plate surface
  3. Streak using an inoculating loop performing back and forth motions from the area previously rolled with the cotton swab to the rest of the agar´s surface
  4. Close the lid and label
  5. Turn the plate upside down
  6. Place on the incubator for 24 hs

MATERIALS

  • Bacterial Solution
  • Agar petri dish
  • Cotton swab
  • Inoculating loop

SPECIAL CONSIDERATIONS

  • When performing the streak motions, movements must be gentle to avoid damaging the agar´s surface.  
  • The plate must be turned upside down to avoid condensation.

POUR PLATE

In the pour plate method, the bacterial solution is mixed on a petri dish with molten agar solution prior to solidification. After incubation, single bacterial colonies will grow on the surface and also through the agar uniformly, allowing to enumerate them. During Antimicrobial Textile Testing this is the preferred method to use when calculating the bacterial concentration of a solution, such as the Single Colony Growth or the bacteria recovered from the samples.

These are the instructions to perform the pour plate technique:

INSTRUCTIONS

  1. Label around the edge of the petri dish
  2. Vortex the bacterial solution
  3. Pipette 1 ml of bacterial solution and place it on a sterile petri dish
  4. Dispense 15 ml of TSA at 50° C onto the petri dish using a serological pipette
  5. Close the lid
  6. Hold the petri dish with two fingers on top of the lid and make figure-eight movements (10 times) to integrate the bacterial solution with the TSA
  7. Partially open the lid and leave inside the hood for 30 minutes to cool down
  8. Close the lid and turn upside down
  9. Place on the incubator for 24 hours

MATERIALS

  • Bacterial solution
  • TSA
  • 25 ml serological pipette
  • Pipette controller
  • 1 ml Pipette tip
  • 1000 ul Pipette
  • Vortexer
  • Petri dish

SPECIAL CONSIDERATIONS

  • The figure-eight motion needs to be performed in a gentle way to prevent the agar from attaching to the lid
  • After the incubation time, when counting the CFU´s, special attention needs to be placed on properly counting the colonies that grow on the surface of the agar as well as the colonies that grow within the agar.